Guillain-Barré syndrome (GBS) is an inflammatory demyelinating disorder of peripheral nerves which occurs a few weeks after a flu-like symptom, and is characterized by rapidly-progressing flaccid-motor paralysis (weakness in muscles of all four limbs), loss of deep tendon reflexes, dysphagia, articulatory disorder, deep sensory disturbance, and vegetative neurosis (cardiac arrhythmia, blood pressure fluctuation). The frequency of Guillain-Barré syndrome is two cases in every 100,000 people per year, and 2,000 to 2,500 people are estimated to be newly affected by the disorder each year across Japan. However, not only is a therapeutic agent capable of achieving complete cure of the disorder undeveloped but also the cause and onset mechanism are yet to be clearly known, which entails the disorder's falling under so-called intractable diseases and being designated a special disease. Recently, plasma-exchange (plasmapheresis) and high-dose intravenous γ-globulin therapy have been reported to be effective treatments for the present syndrome. Further, with respect to the cause of the present syndrome, since autoantibody (anti-lipopolysaccharide antibody) against gangliosides appearing in peripheral nerves, which is associated with acuteness of the symptoms, has been detected in the patients' sera, gangliosides and autoimmune reaction have been pointed out to be closely linking to the onset mechanism. Gangliosides are classified into subclasses according to their molecular structures: GM1, GM2, GD1a, GD1b, GT1a, GQ1b and the like, and respective autoantibodies for each ganglioside are detected in sera of the patients with the disorder. In the aforementioned Guillain-Barré syndrome, it is particularly known that anti-GM1 antibody and anti-GD1a antibody appear as anti-ganglioside antibodies in the serum. Further, an elevated level of anti-GQ1b antibody is almost invariably and specifically observed in the sera in acute phase cases of Guillain-Barré syndrome with ocular muscle paralysis, and in Fisher syndrome cases.
Recently, Campylobacter jejuni, one of the bacteria responsible for foodpoisoning, has been pointed out to be relating to a cause of Guillain-Barré syndrome since there is a molecular homology between the lipopolysaccharide structure of Campylobacter and gangliosides, which may entail the appearance of auto-reactive T cells and B cells. However, it remains unproven whether the appearance of auto-reactive T cells and B cells including autoantibodies leads to any actual organic pathologies. In the animal-experiment level, although a case has been reported where rabbits were immunized with GD1b to develop peripheral neuropathy, there has been no such case where immunizing mice with various types of gangliosides led to their developing any pathological symptoms, which resulted in the absence of such disorder-mouse model suitable for Guillain-Barré syndrome. In cases like the above where rabbits and rats were induced to develop the disorder, the animals exhibited low incidence and mild symptoms which made themselves inappropriate for the model. Further, the absence of suitable model has prevented the development of a therapeutic agent and method.
Likewise, Fisher syndrome is known as a variant of Guillain-Barré syndrome. About 5% of Guillain-Barré syndrome cases are estimated to fall under the aforementioned Fisher syndrome, whose symptoms include external ophthalmoplegia, diplopia, ataxia, loss of tendon reflexes, and facial nerve palsy, with a preceding infection of the upper respiratory tract and the like. The symptoms are identical to those of Guillain-Barré syndrome except that quadriplegia is not induced in humans. Further, in Fisher syndrome cases, elevated level of blood-IgG-antibody titer against ganglioside GQ1b has been reported while as with the case of the above Guillain-Barré syndrome, onset mechanism is unknown and the therapeutic agent is yet to be developed.
The acuteness of these disorders is at its peak about a month after the first symptom appears, gradually getting milder with the recovery period of a few months to a year. The prognosis for the patients with these disorders is relatively good, but the cases with residual deficits are not rare. Moreover, since the patients are forced to experience mental distress, hospitalization and outpatient visits over about a year, the development of the therapeutic agent and treating method for the disorder is ardently awaited by the medical-service community including patients, their families, and physicians.
Meanwhile, on cell surfaces of the immune system and the like, receptors which recognize and bind to Fc part of Ig (hereinafter referred to as “FcR”) are present, among which, Fcγ receptors, the receptor proteins which specifically bind to γ chain of IgG in body fluid (hereinafter referred to as FcγR), are broadly classified into three types based on the gene-structure similarity: Type I (CD64 antigen), Type II (CD32 antigen), and Type III (CD16 antigen). In contrast to other FcRs, FcγRII has low affinity for monomeric IgG and binds to multivalent IgG that has become an immunocomplex, to broadly appear on blood-forming stem cells including monocytes, macrophages, polymorphonuclear leukocytes (PMN), mast cells, blood platelets, some T-cell lymphocytes and some B-cell lymphocytes. Further, there present three types of receptors of FcγRII differing in gene sequences: FcγRIIA, FcγRIIB, and FcγRIIC. All these receptors are mapped to the chromosome 1q23 region.
Unlike other FcRs, the above FcγRIIB possesses an amino-acid sequence which transmits inhibitory signals to intracellular domains without associating with γ chain (ITIM: Immunoreceptor Tyrosine-based Inhibition Motif) (Immunol. Rev. 125, 49-76, 1992, Science 256, 1808-1812, 1992). To elucidate such physiological functions of FcγRIIB, the present inventors have already generated FcγRIIB-deficient mice (Nature 379, 346-349, 1996), as well as arthritis-mouse model which can be obtained by immunizing FcγRIIB-deficient mice with Type II collagen (J. Exp. Med. 189, 187-194, 1999), and autoimmune-disorder animal model (Japanese Laid-Open Patent Application No. Heisei 08-289699).
Previously, appropriate animal model for investigating the onset mechanism of Guillain-Barré syndrome, an inflammatory demyelinating disorder, did not exist. The object of the present invention is to provide non-human animal model that developes Guillain-Barré syndrome (Fisher syndrome) and more specifically, to provide non-human animal model of Guillain-Barré syndrome which can be obtained by immunizing FcγRIIB-gene-deficient non-human animal with ganglioside GQ1b, and a screening method of a therapeutic agent for Guillain-Barré syndrome using the aforementioned non-human animal model.
The present inventors made a keen study to solve the above problem and they endeavored to generate Guillain-Barré syndrome mouse model by using FcγRIIB-gene-deficient mice and immunized them with gangliosides GM1, GM2, GD1a and GQ1b together with Freund's adjuvant every three weeks four times in total. As a result, of those immunized with ganglioside antigen, FcγRIIB-gene-deficient mice immunized with GQ1b exhibited peripheral neuropathy in which paralysis of their tails and hind legs was observed. These mice demonstrated an elevated level of antibody titer against GQ1b, which symptom appeared consistent with that of Fisher syndrome, a variant of Guillain-Barré syndrome, where humans are commonly observed to develop autoantibody against GQ1b. The present inventors have thus completed the present invention by discovering that a new disorder-mouse model can be generated for Guillain-Barré syndrome (Fisher syndrome). The present inventors have also established a screening method of an effective therapeutic agent for the syndrome based on the present invention.